Our aims in this competitive renewal application are shaped by our observations during the previous funding period that confirmed the critical role of cyclooxygenase (COX)-2 in cancer invasion and metastasis. We found that COX-2 was induced in hostile environments and played a critical role in mediating invasion and metastasis. COX-2 expression also increased following treatment with a conventional chemotherapy agent, 5-fluorouracil. Molecular characterization and functional imaging have identified new functional roles for COX-2 that have created new possibilities for more effective COX-2 targeting, and for imaging COX-2 expression and activity. Our data demonstrated the importance of targeting this pathway in cancer, and finding strategies to image COX-2 expression and activity. These data highlighted the importance of expanding our understanding of the role of COX-2 in altering the tumor phenotype to identify additional pathways, networks, and targets that mediate these alterations, and the importance of noninvasively identifying tumors that have increased COX-2 expression, to select for COX-2 targeting. These observations have led us to focus on three new aims that will advance our understanding of the role of inflammation in cancer progression, treatment and metastasis. These studies will be performed using triple negative human breast cancer xenograft models with different COX-2 expression levels. Our purpose in Aim 1 will be to identify the molecular causes of the effect of COX-2 on choline metabolism. These studies can uncover new biomarkers and new targets to use in combination with COX-2 targeting, to achieve improved effectiveness. In Aim 2 we will investigate the role of COX-2 in altering the extracellular matrix structure and function using imaging. In Aim 3 we will develop optical and MRS based probes to report on COX-2 expression and activity that may, in the future, lead to clinically translatable 19F MRS probes to detect COX-2 activity. The studies in this application will advance our insight and identify new strategies to exploit this critically important target in cancer treatment. PUBLIC HEALTH RELEVANCE: COX-2 is a critically important target in cancer that significantly influences a range of characteristics such as angiogenesis, invasion and metastasis. In this application we intend to uncover new targets that interact with COX-2, and identify the effect of COX-2 expression on extracellular matrix structure and function. We also intend to develop probes to noninvasively image COX-2 expression and activity that will allow us to further understand the role of this enzyme in cancer and allow us to effectively target it.